This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Phosphoprotein Phosphatase 1, PP1, is a ubiquitous serine/threonine phosphatase (330 residues, ~ 38 kDa) expressed in all eukaryotes that regulates a wide array of cellular processes. The promiscuity of PP1 is greatly reduced by interaction with several inhibitory and targeting proteins. By interacting with residues of the active site of PP1, inhibitory proteins prevent to access catalytic residues of the enzyme. The targeting proteins relocate PP1 to a particular cellular milieu and interact with the surface of the catalytic core in such a way that only certain binding sites are exposed, whereas, others are sites are sterically occluded. Current crystallographic studies of PP1 in complex with small molecule toxins, inhibitory proteins and targeting proteins have revealed that the surface and conformation of the catalytic core remains invariant;however, all of the proteins studied to date have shown markeedly different themes in their interactions with the PP1. To shed new insights on PP1 regulation by targeting proteins, we have initiated work with one such protein, NIPP (Nuclear Inhibitor of PP1). NIPP targets PP1 after interaction with various proteins implicated in four different cellular processes (cell division, gene silencing, DNA repair and RNA splicing). Since targeting of NIPP1 to PP1 is a major event in cell cycle regulation;thus, we are interested in understanding this relationship at the atomic level. After exhaustive biophysical characterization of the monomeric form of the PP1-binding domain of NIPP, we have moved to studying the holoenzyme complex;to this end, we have determined the Kd with ITC (low nM), used data from complex NMR experiments to optimize samples for crystallization trails and attained two dimensional needle-like crystals.